The evolutionary relationships of twenty-five of the most significant human pathogenic fungi will be determined in relation to fifty of their close higher fungal relatives by applying a powerful new DNA amplification and sequencing technology to study fungal ribosomal RNA genes. The polymerase chain reaction methodology, which has been recently applied to the diagno- sis of viral diseases such as HIV, HTLV-I, and human papilloma virus, will be used to amplify and directly sequence the nuclear small subunit rRNA gene. and other regions of the rDNA repeat, of the pathogens and other fungi. Pathogenic fungi for study include Ascomycetes (e.g., Ajellomyces capsulatus, A. dermatitidis, and Arthroderma species), Basidiomycetes (e.g., Filobasidiella neoformans), and fungi of uncertain placement (e.g., Coccidioides immitis, Candida albicans, and Trichosporon beigelii). The nucleotide sequence variation among these and other pathogens, and other higher fungi, will be used to construct a molecular phylogeny of Ascomycetes and Basidiomycetes. Knowledge of true phylogenetic relationships is essential to predicting the attributes of fungi, including their susceptibility to methods of prevention and control. The results of this study will allow development of fungal-wide and pathogen-specific amplification primers, and pathogen-- specific identification probes which will improve the speed and accuracy of diagnostic tests for the agents of human mycoses. The results will also be broadly applicable to the fields of mycotoxins in food and feed and fungal plant pathology.